title: Generation of glycosylphosphatidylinositol anchor protein-deficient blood cells from human induced pluripotent stem cells.
abstract: PIG-A is an X-linked gene required for the biosynthesis of glycosylphosphatidylinositol (GPI) anchors; thus, PIG-A mutant cells have a deficiency or absence of all GPI-anchored proteins (GPI-APs). Acquired mutations in hematopoietic stem cells result in the disease paroxysmal nocturnal hemoglobinuria, and hypomorphic germline PIG-A mutations lead to severe developmental abnormalities, seizures, and early death. Human induced pluripotent stem cells (iPSCs) can differentiate into cell types derived from all three germ layers, providing a novel developmental system for modeling human diseases. Using PIG-A gene targeting and an inducible PIG-A expression system, we have established, for the first time, a conditional PIG-A knockout model in human iPSCs that allows for the production of GPI-AP-deficient blood cells. PIG-A-null iPSCs were unable to generate hematopoietic cells or any cells expressing the CD34 marker and were defective in generating mesodermal cells expressing KDR/VEGFR2 (kinase insert domain receptor) and CD56 markers. In addition, PIG-A-null iPSCs had a block in embryonic development prior to mesoderm differentiation that appears to be due to defective signaling through bone morphogenetic protein 4. However, early inducible PIG-A transgene expression allowed for the generation of GPI-AP-deficient blood cells. This conditional PIG-A knockout model should be a valuable tool for studying the importance of GPI-APs in hematopoiesis and human development.
authors.affiliation: Division of Hematology, Department of Medicine, School of Medicine, and.
authors.forename: Xuan
authors.initials: X
authors.lastname: Yuan
authors.forename: Evan M
authors.initials: EM
authors.lastname: Braunstein
authors.forename: Zhaohui
authors.initials: Z
authors.lastname: Ye
authors.forename: Cyndi F
authors.initials: CF
authors.lastname: Liu
authors.forename: Guibin
authors.initials: G
authors.lastname: Chen
authors.forename: Jizhong
authors.initials: J
authors.lastname: Zou
authors.forename: Linzhao
authors.initials: L
authors.lastname: Cheng
authors.forename: Robert A
authors.initials: RA
authors.lastname: Brodsky
mesh_terms: D018952:Antigens, CD34 / Q000235:genetics / Q000378:metabolism; D001427:Bacterial Toxins / Q000378:metabolism; D055415:Bone Morphogenetic Protein 4 / Q000235:genetics / Q000378:metabolism; D019002:CD56 Antigen / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation / Q000235:genetics; D002460:Cell Line; D055786:Gene Knockout Techniques; D020868:Gene Silencing; D050172:Genes, X-Linked; D017261:Glycosylphosphatidylinositols / Q000097:blood / Q000172:deficiency* / Q000235:genetics; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000378:metabolism; D006457:Hemoglobinuria, Paroxysmal / Q000097:blood* / Q000235:genetics; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism; D008297:Male; D008565:Membrane Proteins / Q000097:blood / Q000172:deficiency* / Q000235:genetics; D008648:Mesoderm / Q000196:embryology / Q000378:metabolism; D009154:Mutation; D052899:Pore Forming Cytotoxic Proteins / Q000378:metabolism; D012640:Seizures; D019076:Transgenes; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics / Q000378:metabolism
chemical_list: D018952:Antigens, CD34; D001427:Bacterial Toxins; D055415:Bone Morphogenetic Protein 4; D019002:CD56 Antigen; D017261:Glycosylphosphatidylinositols; D008565:Membrane Proteins; C522918:NCAM1 protein, human; D052899:Pore Forming Cytotoxic Proteins; C080086:phosphatidylinositol glycan-class A protein; C107096:proaerolysin; D040301:Vascular Endothelial Growth Factor Receptor-2
keywords: GPI-anchor protein; Hematopoiesis; Human iPSCs; PIG-A gene knockout